Deriving fractional rate of degradation of logistic-exponential (LE) model to evaluate early in vitro fermentation

Water-soluble components of feedstuffs are mainly utilized during the early phase of microbial fermentation, which could be deemed an important determinant of gas production behavior in vitro. Many studies proposed that the fractional rate of degradation (FRD) estimated by fitting gas production curves to mathematical models might be used to characterize the early incubation for in vitro systems. In this study, the mathematical concept of FRD was developed on the basis of the Logistic-Exponential (LE) model, with initial gas volume being zero (LE₀). The FRD of the LE₀ model exhibits a continuous increase from initial (FRD₀) toward final asymptotic value (FRD_F) with longer incubation time. The relationships between the FRD and gas production at incubation times 2, 4, 6, 8, 12 and 24 h were compared for four models, in addition to LE₀, Generalization of the Mitscherlich (GM), c^th order Michaelis–Menten (MM) and Exponential with a discrete LAG (EXP_LAG). A total of 94 in vitro gas curves from four subsets with a wide range of feedstuffs from different laboratories and incubation periods were used for model testing. Results indicated that compared with the GM, MM and EXP_LAG models, the FRD of LE₀ model consistently had stronger correlations with gas production across the four subsets, especially at incubation times 2, 4, 6, 8 and 12 h. Thus, the LE₀ model was deemed to provide a better representation of the early fermentation rates. Furthermore, the FRD₀ also exhibited strong correlations (P < 0.05) with gas production at early incubation times 2, 4, 6 and 8 h across all four subsets. In summary, the FRD of LE₀ model provides an alternative to quantify the rate of early stage incubation, and its initial value could be an important starting parameter of rate.     

A generalized compartmental model to estimate the fibre mass in the ruminoreticulum: 1. Estimating parameters of digestion

Parameters related to the microbial digestion of nutrients in the ruminoreticulum have been estimated by fitting mathematical models to degradation profiles generated from kinetic studies. In the present paper, we propose a generalized compartmental model of digestion (GCMD) based on implicit theoretical concepts and the gamma probability density function to estimate fibre digestion parameters. The proposed model is consistent to a broader compartmental model presented in a companion paper that integrates aspects of fibre digestion and passage. Different versions of the GCMD were generated by increasing the integer order of time dependency of the gamma function. These versions were fitted to 192 published fibre degradation profiles that were obtained using an in vitro fermentation technique. The quality of fit was evaluated based on the frequency of minimum sum of squares of errors (SSE), the number of runs of signs of residuals, and its likelihood probability calculated according to the Akaike's Information Criterion. The likelihood of the proposed model was also compared to a discrete lag time model (DLT), which is commonly used to interpret fibre degradation profiles. The GCMD had superior quality of fit compared to the DLT and was considered more likely in describing 68.75% of the profiles evaluated. Only 9.38% of the degradation profiles that were fitted to the DLT model had a lower SSE. Even though the degradation profiles studied were generated by incubating feed samples up to 96 h, the true asymptotic limit of fibre degradation can only be achieved by long-term fermentations. This fact leads to questioning the uniformity of the potentially digestible fibre fraction and a further approach based on GCMD-type model was used to account for its heterogeneous nature.     

The effect of rabbit age on in vitro caecal fermentation of starch, pectin, xylan, cellulose, compound feed and its fibre

In vitro gas production kinetics of six different substrates, pectin (PEC), xylan (XYL), starch (STA), cellulose (CEL), commercial compound feed (FEED; 201 g crude protein per kg, 155 g crude fibre per kg, 334 g neutral-detergent fibre (NDF) per kg and 190 g acid-detergent fibre (ADF) per kg) and an NDF prepared from commercial compound feed (NDF_FEED) were determined using the caecum contents of weaned rabbits (36 days of age) and of rabbits at slaughter age (78 days of age) as inoculums. The cumulated gas production over 96 h of incubation was modelled with Gompertz model, and the kinetic parameters compared. The total potential gas production (parameter ‘B’ of the Gompertz model) was not affected (P>0.05) by the inoculum source, except with STA, where rabbits at slaughter weight had significantly higher total potential fermentability (314 ml/g dry matter (DM)) than those at weaning age (189 ml/g DM). Intensities of fermentation (maximum fermentation rate; MFR) of PEC (32.2 ml/h) and XYL (24.4 ml/h) were significantly greater in rabbits at weaning, while that of STA (45 ml/h) was significantly lower than at slaughter age (23.0, 14.3 and 14.0 ml/h for PEC, XYL and STA, respectively). The MFRs of CEL and NDF_FEED were very similar between inoculum sources. In the first 10 h of fermentation which correspond to the normal retention time of the substrates in the caecum, the highest amount of gas was produced from PEC, followed by FEED and XYL. These substrates had a time of maximum fermentation rate (TMFR) at both rabbit ages short enough (8.0 and 9.5 h for PEC, 9.5 and 6.6 h for FEED, 13.7 and 14.2 h for XYL at weaning and at slaughter age, respectively) to be almost completely fermented in vivo.     

Intracellular metabolite profiling and the evaluation of metabolite extraction solvents for Clostridium carboxidivorans fermenting carbon monoxide

Clostridium carboxidivorans ferments CO, CO₂, and H₂ via the Wood-Ljungdahl pathway. CO, CO_2, and H₂ are unique substrates, unlike other carbon sources like glucose, so it is necessary to analyze intracellular metabolite profiles for gas fermentation by C. carboxidivorans for metabolic engineering. Moreover, it is necessary to optimize the metabolite extraction solvent specifically for C. carboxidivorans fermenting syngas. In comparison with glucose media, the gas media allowed significant abundance changes of 38 and 34 metabolites in the exponential and stationary phases, respectively. Especially, C. carboxidivorans cultivated in the gas media showed changes of fatty acid metabolism and higher levels of intracellular fatty acid synthesis possibly due to cofactor imbalance and slow metabolism. Meanwhile, the evaluation of extraction solvents revealed the mixture of water-isopropanol-methanol (2:2:5, v/v/v) to be the best extraction solvent, which showed a higher extraction capability and reproducibility than pure methanol, the conventional extraction solvent. This is the first metabolomic study to demonstrate the unique intracellular metabolite profiles of the gas fermentation compared to glucose fermentation, and to evaluate water-isopropanol-methanol as the optimal metabolite extraction solvent for C. carboxidivorans on gas fermentation.     

The Interplay of Proton,Electron, and Metabolite Supply for Photosynthetic H2 Production in Chlamydomonas reinhardtii

To obtain a detailed picture of sulfur deprivation-induced H₂ production in microalgae, metabolome analyses were performed during key time points of the anaerobic H₂ production process of Chlamydomonas reinhardtii. Analyses were performed using gas chromatography coupled to mass spectrometry (GC/MS), two-dimensional gas chromatography combined with time-of-flight mass spectrometry (GCxGC-TOFMS), lipid and starch analysis, and enzymatic determination of fermentative products. The studies were designed to provide a detailed metabolite profile of the solar Bio-H₂ production process. This work reports on the differential analysis of metabolic profiles of the high H₂-producing strain Stm6Glc4 and the wild-type cc406 (WT) before and during the H₂ production phase. Using GCxGC-TOFMS analysis the number of detected peaks increased from 128 peaks, previously detected by GC/MS techniques, to ∼1168. More detailed analysis of the anaerobic H₂ production phase revealed remarkable differences between wild-type and mutant cells in a number of metabolic pathways. Under these physiological conditions the WT produced up to 2.6 times more fatty acids, 2.2 times more neutral lipids, and up to 4 times more fermentation products compared with Stm6Glc4. Based on these results, specific metabolic pathways involving the synthesis of fatty acids, neutral lipids, and fermentation products during anaerobiosis in C. reinhardtii have been identified as potential targets for metabolic engineering to further enhance substrate supply for the hydrogenase(s) in the chloroplast.     

In vitro gas production kinetics and short-chain fatty acid production from rumen incubation of diets supplemented with hop cones (Humulus lupulus L.)

The aim of this study was to assess the effects of hop cones (Humulus lupulus L.) from two varieties Aurora and Dana, differing in their α- and β-acid contents, on rumen microbial activity measured with in vitro gas production kinetics and short-chain fatty acids (SCFA) production. Hop cones were added to the total mixed dairy cow ration (CONT) in concentrations simulating a cow’s daily intake of 50, 100 and 200 g of hop cones – the concentrations of hop cones expressed on a substrate basis were 43, 82 and 153 mg/g of substrate. Substrates were anaerobically incubated in glass syringes, and gas production kinetic parameters were determined by fitting data with the Gompertz model. Gas produced after 24 h (Gas24), maximum fermentation rate (MFR) and time of maximum fermentation rate (TMFR) were calculated from the estimated gas production kinetic parameters. After 24 h of incubation, the fermentation liquids of each substrate were taken for the determination of SCFA. Increasing the hop cone concentration decreased the total potential gas production, Gas24, MFR and shortened TMFR. The highest hop cone concentration significantly decreased acetic and butyric acid productions and total SCFA production after 24 h of incubation, but not propionic acid production, resulting in a decreased ratio between acetic acid and propionic acid.     

Reducing agent can be omitted in the incubation medium of the batch in vitro fermentation model of the pig intestines

Over the past decade, in vitro methods have been developed to study intestinal fermentation in pigs and its influence on the digestive physiology and health. In these methods, ingredients are fermented by a bacterial inoculum diluted in a mineral buffer solution. Generally, a reducing agent such as Na₂S or cysteine-HCl generates the required anaerobic environment by releasing metabolites similar to those produced when protein is fermented, possibly inducing a dysbiosis. An experiment was conducted to study the impact of two reducing agents on results yielded by such in vitro fermentation models. Protein (soybean proteins, casein) and carbohydrate (potato starch, cellulose) ingredients were fermented in vitro by bacteria isolated from fresh feces obtained from three sows in three carbonate-based incubation media differing in reducing agent: (i) Na₂S, (ii) cysteine-HCl and (iii) control with a mere saturation with CO₂ and devoid of reducing agent. The gas production during fermentation was recorded over 72 h. Short-chain fatty acids (SCFA) production after 24 and 72 h and microbial composition of the fermentation broth after 24 h were compared between ingredients and between reducing agents. The fermentation residues after 24 h were also evaluated in terms of cytotoxicity using Caco-2 cell monolayers. Results showed that the effect of the ingredient induced higher differences than the reducing agent. Among the latter, cysteine-HCl induced the strongest differences compared with the control, whereas Na₂S was similar to the control for most parameters. For all ingredients, final gas produced per g of substrate was similar (P>0.10) for the three reducing agents whereas the maximum rate of gas production (R_max) was reduced (P<0.05) when carbohydrate ingredients were fermented with cysteine-HCl in comparison to Na₂S and the control. For all ingredients, total SCFA production was similar (P>0.10) after 24 h of fermentation with Na₂S and in the control without reducing agent. Molar ratios of branched chain-fatty acids were higher (P<0.05) for protein (36.5% and 9.7% for casein and soybean proteins, respectively) than for carbohydrate (<4%) ingredients. Only fermentation residues of casein showed a possible cytotoxic effect regardless of the reducing agent (P<0.05). Concerning the microbial composition of the fermentation broth, most significant differences in phyla and in genera ascribable to the reducing agent were found with potato starch and casein. In conclusion, saturating the incubation media with CO₂ seems sufficient to generate a suitable anaerobic environment for intestinal microbes and the use of a reducing agent can be omitted.     

In vitro fermentation of various carbohydrate-rich feed ingredients combined with chyme from pigs

Increased carbohydrate fermentation, compared with protein fermentation, could benefit gut health. In two in vitro experiments, the effect of carbohydrate-rich feed ingredients on fermentation characteristics of ileal chyme from pigs was assessed, using the cumulative gas production technique. Ingredients of the first experiment included gums, inulins, pectins, transgalacto-oligosaccharides, lactose and xylan. In the second experiment, a gum, pectin and transgalacto-oligosaccharides were added at different starting weights, to determine their effects on fermentation characteristics of chyme, in relation to differences in the carbohydrate concentrations. In comparison to fermentation of chyme alone, added carbohydrates led to higher total gas production (p < 0.05), faster maximum rate of gas production (except for xylan) (p < 0.05), and a decreased branched-chain fatty acids to straight chain fatty acids ratio (BCR) (p < 0.05). In the second experiment, for all carbohydrate ingredients, the BCR decreased with increasing starting weights (p < 0.05). If these supplemented dietary carbohydrates were to reach the terminal ileum of the living animal, carbohydrate fermentation in the large intestine could be stimulated, which is known to have beneficial effects on host health.     

Monitoring the biochemical hydrogen and methane potential of the two-stage dark-fermentative process

A two-step process has been recently proposed whereby the products of biological hydrogen production processes are used as substrates for biological methane production. The aim of the present study is to evaluate a simple bench-scale batch procedure for measuring the biochemical hydrogen and methane potential of organic substances as a two-step simulated process. Glucose fermentation showed an hydrogen and methane recovery (measured as the ratio of electron equivalents recovered as hydrogen and methane and electron equivalents of the initial substrate added) from the initial substrate of 13.3% and 75.5%, respectively, that approximates mass balance closure. On the contrary, gas recoveries ranging from 61% to 75% were measured from wastes originating from the food-industry. Moreover, the results demonstrate that the substrate origins significantly influence the ratio of H₂ and CH₄ recovery.     

“7-in” medium,a simple medium for the rapid presumptive identification of bothSalmonella andShigella

A one-tube method using a homogeneous medium for the rapid screening of colonies forSalmonella as well asShigella is described. The selection is based on the following properties: 1) glucose fermentation (acid, gas), 2) lactose fermentation, 3) saccharose fermentation, 4) production of H₂S, 5) urease activity, 6) motility, and 7) indole production.     

Effects of exogenous enzymes on in vitro gas production kinetics and ruminal fermentation of four fibrous feeds

This study was conducted to investigate effects of increasing doses: 0 (control), 6 (low), 12 (medium) and 24 (high) mg/g DM of ZADO^® enzyme preparation mixture (ENZ) on in vitro gas production (GP) and some ruminal fermentation parameters of the fibrous feeds Saccharum officinarum (leaves), Andropogon gayanus (leaves), Pennisetum purpureum (leaves) and Sorghum vulgare (straw). Rumen liquor was obtained from two Brown Swiss cows fitted with permanent rumen cannulae fed a total mixed ration of a 500:500 commercial concentrate and alfalfa hay ad libitum. The GP was recorded at 2, 4, 6, 8, 10, 12, 24, 48, 72 and 96 h of incubation. After 96 h, the incubation was stopped and the pH of the mixture was determined and filtrate used to determine dry matter degradability (DMD), partitioning factor (PF₉₆), gas yield (GY₂₄), in vitro organic matter digestibility (OMD), metabolizable energy (ME), short chain fatty acids (SCFA), and microbial crude protien production (MCP). In general, the crude protein (CP) content of the fibrous feeds was low and ranged from 23 g/kg DM (S. officinarum) to 44 (A. gayanus). The fibre contents (i.e., NDFom and ADFom) were highest (P<0.05) in S. officinarum. Increasing ENZ dose linearly increased (P<0.05) GP of all fibrous feeds and had a quadratically increased (P<0.05) asymptotic gas production in P. purpureum and S. vulgare and rate of gas production in S. officinarum and S. vulgare. Addition of ENZ also quadratically increased (P<0.05) GP at all incubation times in S. officinarum and S. vulgare, and A. gayanus, but only at 72 h in A. gayanus. The parameters of ruminal fermentation of OMD, ME, GY₂₄ and SCFA linearly increased (P<0.05) and MCP linearly decreased (P<0.05) with the ENZ addition. Addition of enzyme affected ruminal fermentation of our feeds differently, mainly dependent on their fibre content, although dosage of enzyme was also important as impacts generally increased at higher dosages of ENZ.     

Minimization of Glycerol Production during the High-Performance Fed-Batch Ethanolic Fermentation Process in Saccharomyces cerevisiae, Using a Metabolic Model as a Prediction Tool

On the basis of knowledge of the biological role of glycerol in the redox balance of Saccharomyces cerevisiae, a fermentation strategy was defined to reduce the surplus formation of NADH, responsible for glycerol synthesis. A metabolic model was used to predict the operating conditions that would reduce glycerol production during ethanol fermentation. Experimental validation of the simulation results was done by monitoring the inlet substrate feeding during fed-batch S. cerevisiae cultivation in order to maintain the respiratory quotient (RQ) (defined as the CO₂ production to O₂ consumption ratio) value between 4 and 5. Compared to previous fermentations without glucose monitoring, the final glycerol concentration was successfully decreased. Although RQ-controlled fermentation led to a lower maximum specific ethanol production rate, it was possible to reach a high level of ethanol production: 85 g · liter⁻¹ with 1.7 g · liter⁻¹ glycerol in 30 h. We showed here that by using a metabolic model as a tool in prediction, it was possible to reduce glycerol production in a very high-performance ethanolic fermentation process.     

Conversion of Solid Organic Wastes into Oil via Boettcherisca peregrine (Diptera: Sarcophagidae) Larvae and Optimization of Parameters for Biodiesel Production

The feedstocks for biodiesel production are predominantly from edible oils and the high cost of the feedstocks prevents its large scale application. In this study, we evaluated the oil extracted from Boettcherisca peregrine larvae (BPL) grown on solid organic wastes for biodiesel production. The oil contents detected in the BPL converted from swine manure, fermentation residue and the degreased food waste, were 21.7%, 19.5% and 31.1%, respectively. The acid value of the oil is 19.02 mg KOH/g requiring a two-step transesterification process. The optimized process of 12∶1 methanol/oil (mol/mol) with 1.5% H₂SO₄ reacted at 70°C for 120 min resulted in a 90.8% conversion rate of free fatty acid (FFA) by esterification, and a 92.3% conversion rate of triglycerides into esters by alkaline transesterification. Properties of the BPL oil-based biodiesel are within the specifications of ASTM D6751, suggesting that the solid organic waste-grown BPL could be a feasible non-food feedstock for biodiesel production.     

Comparison of bovine rumen liquor and bovine faeces as inoculum for an in vitro gas production technique for evaluating forages

Two experiments were undertaken using the in vitro gas production technique of Theodorou et al. [Anim. Feed Sci. Technol. 48 (1994) 185] to compare rumen liquor (RL) and faeces (FA) as inocula for fermenting gramminaceous forages over 96 h periods. Experiment 1 used 12 forages of differing in vivo digestibility (ammonia treated wheat straw, field-cured hay (Lolium perenne) and 10 artificially dried grasses (L. perenne) harvested at different maturities). Experiment 2 used seven maize-silage based forages (whole plant, stover, leaf, lower stem, middle stem, upper stem and husk). In both experiments, rumen liquor and faeces were obtained from two cows in early lactation, each fed daily with 9.4 kg DM of grass silage and 9.0 kg DM of concentrate. Rumen contents were sampled through the fistula, before morning feeding; faeces were sampled from the rectum, immediately afterwards. Rumen liquor (250 ml) was prepared by straining contents through two layers of muslin, adding the solids after blending with 250 ml of buffer and re-straining. Faeces were prepared by mixing (300 ml) with 150 ml of buffer and straining through two layers of muslin and adding a homogenate of the solids and 150 ml of buffer after straining. Data were fitted to the model of France et al. [J. Theor. Biol. 163 (1993) 99]. All model parameters showed FA to have a poorer fermentation capacity than RL. In both experiments, potential gas production volumes (A) were lower (on average 52.9 ml (18.5%)) and lag times longer (on average 2.9 h) for FA compared to RL. Fractional rate of fermentation at half asymptote (T/2) was generally greater for RL than FA (overall means, 0.042, 0.028) and the time required to T/2 being less (overall means, 21.9, 35.4 h). However, potential gas production (A) was highly correlated between RL and FA: Experiment 1 (r²=0.94, 11 forages, excluding ammonia treated straw) and Experiment 2 (r²=0.83, six forages, excluding middle stem). In Experiment 1, organic matter digestibility in vivo (OMD_IV) was also highly correlated with both OMD_FA (r²=0.77, 11 forages) and OMD_RL (r²=0.89, 11 forages); OMD_RL and OMD_FA were also highly correlated (r²=0.81). Similar correlations occurred in Experiment 2. It is concluded that faeces have potential as an alternative inoculum to rumen liquor for in vitro gas production techniques, but methods of overcoming the longer lag phase with faeces require further research.     

The use of model-fitting in the interpretation of 'dual' uptake isotherms

Abstract. Published data of the concentration dependence of the uptake rate (uptake isotherms) of K⁺ Na⁺, Cl^?, SO2^?₄, and L-lysine in barley roots, and glucose and 3-O-methylglucose in potato tuber tissue, were re-examined. In as much as these isotherms yield non-linear, concave upward Eadie-Hofstee plots, they might have been termed ‘dual’ isotherms. In addition, all these isotherms have been considered to display discontinuous transitions in gradient. The following models that yield continuous isotherms were fitted to the isotherms: (1) the sum of a Michaelis-Menten term and a linear term; (2) the sum of two Michaelis-Menten terms; (3) the sum of two Michaelis-Menten terms and a linear one. Goodness of fit was judged from: (i) the weighted mean square of deviates; (ii) the standard errors of the kinetic parameters; (iii) the algebraic significance of the terms; (iv) a Rankits plot of the residuals; (v) a Runs test on the residuals. For the precise and detailed isotherms of SO^2? uptake, only model (3) gave a fit that was satisfactory in all respects. There appeared to be no reason to consider these isotherms as multiphasic. The same conclusion was reached for the L-lysine uptake isotherms. For the other isotherms the results were less conclusive. Thai for K⁺ and Na+ could, at any rate, be described satisfactorily by a continuous model, the best fit being obtained with model (2). The uptake isotherms of Cl^? and 3-0-methylglucose could be best described by model (2), and that of glucose by model (3), only the result of the Runs test being unsatisfactory. It is concluded that there is hardly any evidence that the presumed ‘jumps’ or discontinuities or inflections in the gradient of uptake isotherms are not due to experimental error in the data. It is suggested that many uptake isotherms may be described by model (3), although the reason for this is still incompletely understood.     

Influence of protein fermentation on gas production profiles

With modern equipment, accurate gas-production profiles can be obtained reflecting the organic-matter fermentation in rumen fluid. Although the gas production caused by fermentation of carbohydrates is well understood and described, ignoring the influence of protein fermentation may lead to misinterpretation of the gas-production data. Gas-production profiles, from grass samples differing in growing days, and hence in protein content, showed an unexpected low gas production for the young samples compared to the old ones. The influence of protein fermentation on gas-production profiles was studied by incubation of mixtures of casein with glucose, Zulkovsky starch and/or potato starch. After prolonged incubation, the fermentation of casein produced only 32% gas compared with carbohydrates and it was calculated that each percentage of protein caused a reduction in gas production of 2.48 ml g⁻¹ organic matter. Relative to potato starch, casein was fermented in an earlier stage of incubation.After correction for the influence of protein fermentation, gas production of the youngest grass sample was the highest and of the oldest sample the lowest. It showed that protein fermentation influenced gas production mainly in the initial hours of incubation, because the major part of protein is part of the soluble fraction. It is concluded that a comparison of gas-production profiles of feed samples differing largely in protein content may lead to a misinterpretation, which necessitates correction for protein fermentation.     

In vitro fermentation of various carbohydrate-rich feed ingredients combined with chyme from pigs

Increased carbohydrate fermentation, compared with protein fermentation, could benefit gut health. In two in vitro experiments, the effect of carbohydrate-rich feed ingredients on fermentation characteristics of ileal chyme from pigs was assessed, using the cumulative gas production technique. Ingredients of the first experiment included gums, inulins, pectins, transgalacto-oligosaccharides, lactose and xylan. In the second experiment, a gum, pectin and transgalacto-oligosaccharides were added at different starting weights, to determine their effects on fermentation characteristics of chyme, in relation to differences in the carbohydrate concentrations. In comparison to fermentation of chyme alone, added carbohydrates led to higher total gas production (p < 0.05), faster maximum rate of gas production (except for xylan) (p < 0.05), and a decreased branched-chain fatty acids to straight chain fatty acids ratio (BCR) (p < 0.05). In the second experiment, for all carbohydrate ingredients, the BCR decreased with increasing starting weights (p < 0.05). If these supplemented dietary carbohydrates were to reach the terminal ileum of the living animal, carbohydrate fermentation in the large intestine could be stimulated, which is known to have beneficial effects on host health.     

Coupling of microbial kinetics and oxygen transfer for analysis and optimization of gluconic acid production with Aspergillus niger

Gluconic acid fermentation has been widely used for the analysis of various aspects of kinetics and gas liquid transfer of oxygen. Most of these studies are, however, restricted to processes with bacteria. Mathematical models for industrially important productions with fungi have not been elaborated. In the experimental part of this work computer coupled fermentations of gluconic acid production with Aspergillus niger NRRL 3 have been performed. Knowledge of the stoichiometric relationship in the key reaction (glucose oxidase) provides an excellent opportunity for on-line estimation of glucose, biomass and product gluconate from oxygen uptake and carbon dioxide evolution rates. Starting then from experimental observations on the pH-depending oxygen kinetics of gluconic acid formation and influences of product concentrations on the growth of Aspergillus niger a mathematical framework is developed in which the kinetics of growth and production are coupled with gas liquid oxygen transfer. The model can be successfully applied to simulations of the experimental results of gluconic acid fermentations with cyclic addition of glucose. An important aspect in the coupling of transport and microbial reaction in this model is the incorporation of the influence of sugar and gluconate on the solubility of oxygen and k _La via changes of viscosities and molecular diffusivities. With the development of such a comprehensive model, it appears feasible to investigate the influence of various process conditions (sugar feeding, pressure, optimal pH profiles) and to study their possible impacts on the productivity of the overall process.     

Modeling alcoholic fermentation of glucose/xylose mixtures by ethanologenic Escherichia coli as a function of pH

In order to understand the effect of pH on growth and ethanol production in ethanologenic Escherichia coli, we investigated the kinetic behavior of ethanologenic E. coli during alcoholic fermentation of glucose or xylose in a controlled pH environment and the fermentation of glucose, xylose, or their mixtures without pH control. Based on the Monod equation, an unstructured and unsegregated kinetic model was proposed as a function of the pH of the fermentation medium. The pH effects on cell growth, sugar consumption, and ethanol production were taken into account in the proposed model. Both cell growth and ethanol production were found to be significantly influenced by the pH of the fermentation medium. The optimal pH range for ethanol production by ethanologenic E. coli on either glucose or xylose was 6.0–6.5. The highest value of the maximum specific growth rate (μ _m) was obtained at pH 7.0. In the kinetic model of the fermentations of the sugar mixture, two inhibition terms related to glucose concentrations were included in both the cell growth and ethanol production equations because of the strong inhibitions of glucose and glucose metabolites on xylose metabolism. A good fit was found between model predictions and experimental data for both single-sugar and mixed-sugar fermentations without pH control within the experimental domain.     

Actinomycetes scale‐up for the production of the antibacterial,nocathiacin

An Amycolatopsis fastidiosa culture, which produces the nocathiacin class of antibacterial compounds, was scaled up to the 15,000 L working volume. Lower volume pilot fermentations (600, 900, and 1,500 L scale) were conducted to determine process feasibility at the 15,000 L scale. The effects of inoculum volume, impeller tip speed, volumetric gas flow rate, superficial gas velocity, backpressure, and sterilization heat stress were examined to determine optimal scale‐up operating conditions. Inoculum volume (6 vs. 2 vol %) and medium sterilization (R_o of 68 vs. 92 min^?1) had no effect on productivity or titer, and higher impeller tip speeds (2.1 vs. 2.9 m/s) had a slight effect (20% decrease). In contrast, higher backpressure, incorporating increased head pressure at the 15,000 L scale (1.2 vs. 0.7 kg/cm²) and low gas flow rates (0.25 vs. 0.8 vvm), appeared to be problematic (40–50% decrease). High off‐gas CO₂ levels were likely reasons for observed lower productivity. Consequently, air flow rate for this 25‐fold scale‐up (600–15,000 L) was controlled to match off‐gas CO₂ profiles of acceptable smaller scale batches to maintain levels below 0.5%. The 15,000 L‐scale fermentation achieved an expected nocathiacin I titer of 310 mg/L after 7 days. Other on‐line data (i.e., pH, oxygen uptake rate, and CO₂ evolution rate) and off‐line data (i.e., analog production, glucose utilization, ammonium production, and dry cell weight) at the 15,000 L scale also tracked similarly to the smaller scale, demonstrating successful fermentation scale‐up. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2009